U46619

Immortalized tubular epithelial cells (HK-2) were cultured in DMEM/F12 medium supplemented with 10% FBS. After synchronization, cells were treated with 20% FSGS patients’ serum (PS) or 40 nM C3a (204881, Merck-Calbiochem). For intervention studies, 1 μM of C3aR antagonist SB290157 (sc-222291, Santa Cruz), 100 μg/ml of eculizumab (Soliris, Alexion Pharmaceuticals), 10 μM of p38 MAPK inhibitor SB203580 (sc-3533, Santa Cruz), 50 μM of ERK inhibitor PD98059 (sc-3532, Santa Cruz), 10 μM of Akt inhibitor MK2206 (sc-364537, Santa Cruz) or 1 μM of U-46619 (sc-201242, Santa Cruz) was given 30 min before treatments. To infect HK-2 cells with plenti-CMV-LOC105375913, plenti-CMV-snail or plenti-CMV-XBP-1s plasmid, the lentiviral stock was mixed with polybrene (1 µg/ml) and added to cells. C/EBPβ siRNA (sc-44251), Elk-1 siRNA (sc-35290), ERα siRNA (sc-29305), GR siRNA (sc-35505), snail siRNA(sc-38398) and XBP-1s siRNA (sc-38627) were bought from Santa Cruz (Dallas, Texas, USA). LOC105375913 siRNA was bought from Thermo fisher (4390771, Carlsbad, California, USA). Transfection of siRNA, miRNA mimics or miRNA antisense oligonucleotide (ASO) was conducted with Lipofectamine 2000.

Fasudil hydrochloride was purchased from FUJIFILM Wako Pure Chemical (Osaka, Japan). U46619, a thromboxane A2 (TXA2) receptor agonist, was purchased from Santa Cruz Biotechnology (Texas, USA). These compounds were administered into both luminal (upper) and abluminal side (lower compartment) of the cell culture inserts.

Poly-L-lysine (PLL) (10x) was purchased from Beijing Solarbio Science and Technology Co., Ltd. (Beijing, China). DMEM/F12 (1 : 1), Neurobasal Plus Medium, B27 Plus Supplement (50x), HBSS (1x), FBS, and 0.25% trypsin were obtained from Gibco (Grand Island, New York, USA). The Aβ_25-35 fragment and dimethyl sulfoxide (DMSO) were purchased from Sigma (St. Louis, MO, USA). Donepezil hydrochloride, SB203580, and U46619 were obtained from Santa Cruz Biotechnology Inc. (Delaware, USA). L-Glutamine was obtained from Thermo Fisher (Massachusetts, USA). The rabbit polyclonal antibodies against MAP2 were purchased from Proteintech Group, Inc. (Chicago, USA). Rabbit monoclonal antibodies against p38 MAPK, p-p38 MAPK, caspase-3, cleaved caspase-3, and Bax were purchased from Cell Signaling Technology (Beverly, MA, USA). The primary antibody against Bcl-2 was purchased from Abcam (Cambridge, UK). Horseradish peroxidase- (HRP-) conjugated IgG secondary antibody was purchased from Beijing Bioss Biotechnology Co., Ltd. (Beijing, China), and the BCA protein assay kit was obtained from Beyotime Biotechnological Technology Co., Ltd. (Shanghai, China).